Chemistry: natural resins or derivatives; peptides or proteins; – Proteins – i.e. – more than 100 amino acid residues – Separation or purification
Patent
1990-09-04
1993-03-02
Cashion, Jr., Merrell C.
Chemistry: natural resins or derivatives; peptides or proteins;
Proteins, i.e., more than 100 amino acid residues
Separation or purification
435174, 435177, 435243, 435261, 436532, 436538, 436541, C07K 320, C07K 1700, C12N 1100
Patent
active
051910683
DESCRIPTION:
BRIEF SUMMARY
FIELD OF THE INVENTION
The invention relates to a method of removing an antigenic substance from a fluid. As used herein, the term antigenic substance refers broadly to substances to which antibodies can be produced.
DESCRIPTION OF RELATED ART
Affinity separation and purification techniques are known in which an immobilised antibody is used to selectively remove an antigenic substance from a biological fluid. For example, U.K. Patent Application No. 2 135 676 describes a process for producing highly pure erythropoietin by antibody affinity chromatography.
The separation and purification of cell sub-types has been carried out using an antibody affinity chromatography process. In such a process, the antigenic substance is a surface component of the cells to be removed.
A problem associated with the prior art processes is that each specific separation requires the immobilisation of a different antibody. This presents obvious difficulties for the user as well as the supplier of separation equipment and materials since optimal immobilisation conditions vary for each antibody. In addition, immobilisation of small amounts of antibody is costly since immobilisation efficiency usually depends on the antibody concentration used.
The present invention is designed to solve the problem described above by providing a method in which only a single antibody need be immobilised for a range of applications thus reducing unit cost and enabling optimisation. In particular, a single type of separation material and equipment can be used for a range of immunoaffinity separations.
SUMMARY OF THE INVENTION
The invention provides a method of removing an antigenic substance from a fluid which method comprises antigenic substance, and said second antibody being immobilised on a solid phase carrier, and
BRIEF DESCRIPTION OF THE DRAWINGS
The invention is illustrated, by way of example, in the accompanying schematic drawings wherein:
FIG. 1 is a cross sectional view of a preferred solid phase carrier element for use in the invention;
FIG. 2 is a perspective view of the exterior of an apparatus in which the invention may be performed;
FIG. 3 is a longitudinal sectional view taken along line 2--2 of FIG. 2; and,
FIG. 4 is a transverse sectional view taken along lines 3--3 of FIG. 2.
DESCRIPTION OF THE PREFERRED EMBODIMENTS
In a preferred embodiment of the invention the fluid is contacted with the first antibody to form an antibody:antigen binary complex with the antigenic substance and is subsequently contacted with the immobilised second antibody to form the ternary complex.
It is particularly preferred that the first antibody is a mouse antibody and the second antibody is an anti-mouse kappa chain antibody. Since the immobilised antibody binds the kappa chains of mouse antibodies specifically, it interacts with a high proportion of mouse antibodies (approximately 95% of mouse antibodies have a common kappa chain). This is particularly useful since the majority of monoclonal antibodies commercially available is of murine origin.
Preferably, the first antibody and/or the second antibody is a monoclonal antibody.
The method of the invention may be used in respect of a wide variety of antigenic substances. Examples of such substances include haptens, hormones, allergens, viruses, bacteria, toxins, some drugs and proteins.
A particular application of the invention is the separation and purification of cell sub-types e.g. T-cells. For example, a cell population containing a sub-population of cells of interest is exposed to an excess of a kappa chain-containing mouse monoclonal antibody specific for a particular cell surface protein which is only present in that sub-population. After separating the cells from the suspension which contains excess mouse antibody, the cells are contacted with an anti-mouse kappa chain antibody immobilised on a solid phase carrier. Only the cells binding the specific mouse antibody are retained by the solid phase carrier. Since most of the monoclonal antibodies available for cell typing are produced i
REFERENCES:
patent: 5009997 (1991-04-01), Shah et al.
patent: 5081030 (1992-01-01), Civin
Darnell et al., Molecular Cell Biology: Antibodies, (Scientific American Books), pp. 77, 584-586 (1986).
Johnstone et al., Immunochemistry in Practice, (Blockwell Scientific Publications), pp. 227-228 (1985).
Ware et al., J. Immunol. Meth., vol. 74 (1) pp. 93-104 (1984).
O'Grady et al., Chem. Abs. vol. 105 entry #170267w (1986).
Schrader et al. J. Immunol. Meth., vol. 93 (1) pp. 45-53 (1986) (Abstract).
Dean et al., Affinity Chromatography-A Practical Approach, (IRG Press, Washington, DC), pp. 191-206 (1985).
Clark Stephen E.
Daiss John L.
Stickley Frances L.
Thomson Alan R.
Cashion Jr. Merrell C.
Eastman Kodak Company
Rozycki Andrew G.
Wells Doreen M.
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